BackgroundThere are several reports suggesting that hyperosmolarity induces inflammation. We recently showed that Dextran Sodium Sulfate causes inflammatory bowel disease due to hyperosmolarity. The aim of this study was to confirm the link between hyperosmolarity and inflammation by assessing osmolarity values in vivo during inflammation, compare the inflammatory potential of different osmotic agents and finally study the long-term consequences of hyperosmolarity on cell fate.MethodsOsmotic pressures were measured in inflammatory liquids withdrawn from mice subjected to inflammation caused either by subcutaneous injection of Bacille Calmette-Guérin (BCG) or Freund adjuvant. Three epithelial cell lines (HT29, T24 and A549) were exposed up to 48 hours to increasing osmolarities (300, 600, 900 mOsm) of chemically inert molecules such as Mannitol, Propylene Glycol, and Glycerol and inflammatory response was assessed by Enzyme Linked ImmunoSorbent Assay (ELISA) and RNA Protection Assay (RPA). Finally, normal mouse macrophages were exposed to hyperosmotic conditions for long-term culture.ResultsThe inflammation caused either by BCG or Freund adjuvant is correlated to hyperosmolarity in inflammatory liquids. The exposure of cells to the different compounds, whatever their molecular weight, has no effect on the secretion of cytokines as long as the osmolarity is below a threshold of 300 mOsm. Higher osmolarities result in the secretion of proinflammatory cytokines (Interleukin-8, Interleukin-6, Interleukin-1β and Tumor Necrosis factor-α). Long-term hyperosmotic culture extends normal macrophage half-life, from 44 days to 102 days, and alters the expression of p53, Bcl-2 and Bax.ConclusionThe present study further suggests inflammation and hyperosmolarity are closely related phenomena if not synonymous.

Hyperosmolarity can induce pro-inflammatory cytokine responses in colorectal and bladder epithelial cells. Inflammation appears to be the simple consequence of a shift of methylation of PP2A which in turn activates NF-kappaB.

There are several reports suggesting hyperosmotic contents in the feces of patients suffering from inflammatory bowel disease (IBD). Previous works have documented that hyperosmolarity can cause inflammation attributable to methylation of the catalytic subunit of protein phosphatase 2A (PP2A) and subsequent NF-kappaB activation resulting in cytokine secretion. In this study, we demonstrate that dextran sulfate sodium (DSS) induces colitis due to hyperosmolarity and subsequent PP2A activation. Mice were randomized and fed with increased concentrations of DSS (0 mOsm, 175 mOsm, 300 mOsm, and 627 mOsm) for a duration of 3 wk or with hyperosmotic concentrations of DSS (627 mOsm) or mannitol (450 mOsm) for a duration of 12 wk. Long-term oral administration of hyposmotic DSS or mannitol had no demonstrable effect. Hyperosmotic DSS or mannitol produced a significant increase in colonic inflammation, as well as an increase in the weight of sacral lymph nodes and in serum amyloid A protein levels. Similar results were obtained through the ingestion of comparable osmolarities of mannitol. Hyperosmolarity induces the methylation of PP2A, nuclear p65 NF-kappaB activation. and cytokine secretion. The rectal instillation of okadaic acid, a well-known PP2A inhibitor, reverses the IBD. Short inhibiting RNAs (siRNAs) targeted toward PP2Ac reverse the effect of hyperosmotic DSS. The present study strongly suggests that DSS-induced chronic colitis is a consequence of the methylation of PP2Ac induced by hyperosmolarity.

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